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The phosphotyrosyl phosphatase activator, Ncs1p (Rrd1p), functions with Cla4p to regulate the G(2)/M transition in

D A Mitchell1, G F Sprague

  • 1Institute of Molecular Biology, University of Oregon, Eugene, Oregon 97403-1229, USA.

Molecular and Cellular Biology
|January 3, 2001
PubMed

Insights

Investigating essential shared functions of Saccharomyces cerevisiae kinases Ste20p and Cla4p revealed NCS1, a gene crucial for cell cycle progression. Ncs1p interacts with the phosphatase Sit4p, highlighting a novel regulatory pathway essential for cell viability.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Genetics

Background:

  • Saccharomyces cerevisiae kinases Ste20p and Cla4p possess individual and shared essential functions.
  • A strain lacking both Ste20p and Cla4p is inviable, indicating functional overlap.
  • Identifying mutations lethal in the absence of CLA4 (NCS mutations) can elucidate these shared functions.

Purpose of the Study:

  • To investigate the shared essential functions between Saccharomyces cerevisiae p21-activated kinases Ste20p and Cla4p.
  • To identify novel genes involved in essential cellular processes through a genetic screen for mutations lethal in the absence of CLA4.
  • To characterize the function of NCS1/RRD1 and its role in cell cycle regulation and interaction with protein phosphatases.

Main Methods:

  • Genetic screen for mutations lethal in the absence of CLA4 (NCS mutations).
  • Analysis of double mutant phenotypes, including septin localization, cell morphology, and cell cycle progression.
  • Biochemical assays to detect protein complexes (Cla4p-Cdc28p) and protein-phosphatase interactions (Ncs1p-Sit4p).

Main Results:

  • Identified at least 10 complementation groups of NCS mutations.
  • The NCS1/RRD1 gene was characterized, and its absence in combination with cla4Delta resulted in cell cycle arrest with elongated buds and short mitotic spindles.
  • NCS1 encodes a protein similar to mammalian phosphotyrosyl phosphatase activators (PTPAs) and interacts with the yeast phosphatase Sit4p.
  • Genetic interactions (synthetic lethality with SIT4) and biochemical evidence confirmed Ncs1p's role in targeting Sit4p.
  • Loss of both NCS1 and its paralog NOH1 resulted in inviability and G1 arrest, indicating a requirement for PTPA function in G1 progression.

Conclusions:

  • NCS1 plays a critical role in cell cycle progression, likely through its interaction with the Sit4p phosphatase.
  • The Ncs1p-Sit4p complex is essential for proper cell division and viability in Saccharomyces cerevisiae.
  • This study uncovers a novel regulatory pathway involving PTPA activity in cell cycle control, particularly in G1 phase.

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